Crude extract of Ruellia tuberosa L. flower induces intracellular ROS, promotes DNA damage and apoptosis in Triple Negative Breast Cancer Cells

Ethnophamacological relevanceIn the traditional folklore medicine system, the primary uses of Ruellia tuberosa L. include as a diuretic, anti-hypertensive, antipyretic, anti-diabetic, analgesic, and gastroprotective agent. Some reports also demonstrated that it has been used to treat gonorrhea-like diseases. PurposeExploring the anti-cancer potential of the methanolic extract of Ruellia tuberosa L. flower (RTME) with special emphasis on human triple-negative breast cancer (TNBC) and investigating the possible signaling networks and regulatory pathways underlying it. MethodsPreparation of RTME and identifying the possible phytochemicals through GC-MS analysis. The anti-cancer potential of RTME was executed through in-vitro cytotoxicity assay, clonogenic assay, wound healing assay, ROS generation assay, cell cycle arrest, apoptotic nuclear morphology study, cellular apoptosis study, mitochondrial membrane potential (MMP) alteration study, protein and gene expressions alteration study. Apart from this, toxicological status and in-silico molecular docking studies were also conducted. ResultsFrom this study, it was obvious that several phytochemicals within RTME have the potential to act as anti-cancerous agents. RTME was found to exhibit significant in-vitro cytotoxicity along with a reduction in colony formation and inhibition of cell migratory potential in MDA-MB-231 cells. RTME also induced intracellular ROS, promoted G0/G1 cell cycle arrest, caused mitochondrial membrane potential (MMP) alteration, and promoted cell death. From the pro- and anti-apoptotic marker study through the western blot and the qRT-PCR analysis, it was revealed that RTME promoted the intrinsic pathway of apoptosis. Furthermore, blood parameters and histological analysis revealed that RTME doesnt exhibit any toxic effect on female Balb/C mice. Finally, an in-silico molecular docking study revealed that the three identified lead phytochemicals in RTME show strong receptor-ligand interactions with the anti-apoptotic Bcl-2 and give a clue to the possible molecular mechanism of the RTME extract. ConclusionFrom the findings, it was concluded that RTME has a significant therapeutic potential against TNBC which could be an alternative option for anti-cancer drug development. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=188 SRC="FIGDIR/small/586749v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@72780corg.highwire.dtl.DTLVardef@1276029org.highwire.dtl.DTLVardef@1a2d91org.highwire.dtl.DTLVardef@a5cb44_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIPreparation of methanolic extract of Ruellia tuberosa L. flower, C_LIO_LIIdentification of phytochemicals from the methanolic extract of Ruellia tuberosa L. flower, C_LIO_LIMethanolic extract of Ruellia tuberosa L. (RTME) flower exhibited significant anti-cancer potential in triple-negative breast cancer (TNBC) cells, MDA-MB-231 through induction of intracellular ROS, G0/G1 cell cycle arrest, and apoptosis, C_LIO_LIToxicological assessments of RTME on female Balb/C mice, C_LIO_LIIn-silico assessments of lead phytochemicals with the target anti-apoptotic protein, bcl-2 C_LI